Biochemical characteristics of endonuclease activity within the nucleotidyl DNA excision repair pathway of permeable human fibroblasts.
DNA strand breage in response to damage produced by UV (254 nm) radiation was characterized after permeabilization of diploid normal and xeroderma pigmentosum variant fibroblasts. The breakage reaction required ATP, Mg2+ and sucrose for maximal activity and was inhibited by 150 mM Na+ or K+ and 1 mM N-ethylmaleimide. ATP-dependent strand breakage was saturated at UV fluences of above 10 J/m2 and in the presence of DNA precursors breakage was rapidly followed by DNA polymerase and ligase activities to seal the strand breaks. The biochemical features of strand breakage in irradiated permeable cells suggest an enzymatic process. These results, therefore, provide an indication of the biochemical requirements for the rate-limiting strand incision step within the nucleotidyl DNA excision repair pathway.
Kaufmann
W K
WK
Department of Pathology, University of North Carolina, Chapel Hill 27599-7295.
Briley
L P
LP
D000255:Adenosine Triphosphate / Q000494:pharmacology; D002460:Cell Line; D002463:Cell Membrane Permeability; D004247:DNA / Q000528:radiation effects*; D004249:DNA Damage; D011088:DNA Ligases / Q000378:metabolism; D004260:DNA Repair; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D004706:Endodeoxyribonucleases / Q000378:metabolism*; D005033:Ethylmaleimide / Q000494:pharmacology; D005347:Fibroblasts / Q000201:enzymology; D006801:Humans; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D009097:Multienzyme Complexes / Q000378:metabolism*; D009699:N-Glycosyl Hydrolases / Q000378:metabolism*; D011188:Potassium / Q000494:pharmacology; D012964:Sodium / Q000494:pharmacology; D014466:Ultraviolet Rays; D014983:Xeroderma Pigmentosum
D009097:Multienzyme Complexes; C032219:UV endonuclease; D000255:Adenosine Triphosphate; D004247:DNA; D012964:Sodium; D004259:DNA-Directed DNA Polymerase; D004706:Endodeoxyribonucleases; D009699:N-Glycosyl Hydrolases; D011088:DNA Ligases; D008274:Magnesium; D005033:Ethylmaleimide; D011188:Potassium